Project Summary: Recent studies have demonstrated that beneficial and pathogenic microbial associations often use the same host and bacterial surface molecules and bioactive compounds to mediate their interactions at epithelial cell surfaces. These findings beg the question: how does the host promote health by simultaneously discouraging pathogens and encouraging beneficial microbial partners? The symbiosis between the epithelial tissues of the squid Euprymna scolopes and its Gram- negative bacterial partner Vibrio fischeri is being exploited as an experimental system by which to address this critical question. In this association, derivatives of lipopolysaccharide and peptidoglycan of the bacterial partner interact with epithelial cells to direct host development during the establishment of the symbiosis. The proposed research seeks to define the molecular underpinnings of this communication. The principal aims are: 1. To characterize when and where host pattern-recognition receptors for bacterial products are expressed during the onset of the symbiosis - Sensitive techniques to localize specific mRNAs and proteins in animal tissues will be used to ask: How does the expression of host receptors mediate or restrict the interaction of the partners to promote a beneficial association? 2. To determine the influence on symbiosis of defects in V. fischeri's capacity to produce bioactive ligands - Using bacterial genetic techniques developed for V. fischeri, under- and overproducing mutants will be created to ask: How does a normal level of bacterial ligand synthesis promote beneficial interactions? 3. To identify host responses to bacterial ligands using host genomic analyses - Microarray studies of genes of the host tissues expressed in early development will address: What gene-expression patterns of the host underlie the interactions with a beneficial partner? Relevance: Host-receptor/bacterial-ligand interactions have been conserved throughout animal evolution. The binary squid-vibrio symbiosis offers a relatively simple model in which to identify and define the basic, shared mechanisms by which essential bacteria colonize human and other animal epithelia. It provides a powerful complement to the similar, yet more complex, germ-free vertebrate models of beneficial associations.